~70% composed of 8 α-helixes; remainder composed of turns and loops, forming a compact globular shape (much larger when denatured)
Overall shape is devoid of symmetry
Interior contains non-polar residues
except for two histidines to bind iron and oxygen
Exterior contains both polar and non-polar residues
Hydrophobicity and Hydrophilicity
Water soluble proteins are amphipathic – meaning both polar and non-polar
Generally fold in compact structures with non-polar cores
Hydrophilic –R groups (polar) --> Outside
Hydrophobic –R groups (non-polar) --> Inside
Backbone made hydrophobic by hydrogen bonding within α-helix and β-pleated sheets
Polar –R groups made hydrophobic by van der Waals interactions.
Exception: Porin within outer membrane of bacteria is hydrophobic outside (interact with lipid membrane) and hydrophilic inside (allows polar molecules through)
Protein Domains
Two or more compact globular protein regions connected by flexible segment of polypeptide chain
Protein Domains are within the same protein; do not confuse with quaternary structure which involves more than one protein
Quaternary Structure
Spatial arrangement of proteins containing more than one polypeptide chain
Each protein in a quaternary structure is called a subunit
Dimmers contain two identical subunits
Tetramer consists of four subunits
Protein Folding
Denature protein using:
β-mercaptoethanol reduces covalent disulfide bonds to sulfhydryls (-S-S- to –SH + HS-)
Strong reducing agent
Urea and guanidinum chloride can disrupt hydrogen bonds and hydrophobic interactions
Denaturing protein using β-mercaptoethanol, urea and guanidinum chloride destroys enzymatic activity
Protein activity can be recovered when β-mercaptoethanol, urea and guanidinum chloride were removed and oxygen added suggesting correct refolding of the protein with correct disulfide bonds reforming
if only β-mercaptoethanol removed but keep urea and guanidinum chloride, protein can reform but disulfide bonds form randomly and is enzymatically inactive
Proteins have inherent ability to form its correct tertiary structure
“If you leave it alone, it’ll figure out what its supposed to do”
Primary structure specifies conformation of the final tertiary protein
Protein folding in tertiary structure is not a random processes
Levinthal’s paradox – calculated versus actual time for a protein to fold correctly
Predict that it would take 1.6 x 10 e27 years to fold correctly by random search
Actually, E. coli produces 100 active proteins in 5 seconds
Cooperative transition in protein folding – “all or none”
Partially denatured solution has 50% folded and 50% unfolded proteins
Sharp transition between folding native protein and denatured unfolding protein (and vice versa)
Because unfolding proteins form aggregates and takes up space, it is important for proteins to have an “all or none” approach to protein folding to avoid forming aggregates
Chaperones help proteins to correctly fold and avoid aggregates
Cumulative selection aids protein folding
Retention of partially correct intermediates by differences in free energy and leads to increasing stability as the entire correct conformation is achieved
That is, as soon as the first fold is correct, it helps the next fold being correct
Prediction of 3-D protein structures
ab initio predictions
Predict secondary and tertiary structures based by amino acid sequence without reference to known protein structures
Computers used to determine differences in free energy of protein structures
Utility limited by sheer number of possible configurations
Knowledge-based methods
Amino acid sequence of unknown protein compared for sequence compatibility with proteins of known conformation by computer analysis
If significant match is found, it is used as an initial model
Predictions of secondary structures with 6 or fewer residues are 60-70% accurate
Conformational preferences of amino acids are not absolute
Tertiary interactions and microenvironment also play roles not fully understood
Human Disease
Several neurogenerative diseases are disease of protein misfolding.
Infectious protein replicates by transferring protein misfolding in the absence of nucleic acid from α-helices to form insoluble aggregates of β-pleated sheets
Prions are responsible for transmissible spongiform encephalopathies (TSE)
Fatal neurodegenerative disorders affecting humans and other mammals
Introduction to Proteins III Part 1
Dr. Kandace Williams, Ph.D.
Table of Contents
3-D Structures
Tertiary Structure
Myoglobin
Hydrophobicity and Hydrophilicity
Protein Domains
Quaternary Structure
Protein Folding
Prediction of 3-D protein structures
Human Disease
Objectives